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a Novartis company

Comparability assessment in the development of follow-on protein products: Case studies from Omnitrope®

Andreas Premstaller, Ph.D.Biopharmaceutical OperationsSan Francisco, January 14, 2009

2 Comparability assessment in the development of follow-on protein products: Case studies from Omnitrope®/ AP / 2009.01.14

Outline

• Analytical tools for the characterization of somatropin• Comparability studies in the development of Omnitrope®:

Scope, approaches and execution• Case studies of comparability assessments:

− Biosimilarity− Process changes− Novel formulations

Analytical toolsComprehensive characterization of

somatropin


• Recombinant human growth hormone(rhGH) from modified E. coli

• Chemically identical to pituitary growth hormone, non-glycosylated protein

• 1 chain of 191 amino acids, 2 disulfide bridges, with 4 antiparallel α-helices

• C990H1528N262O300S7

• Mass 22125

Somatropin, the active ingredient of Omnitrope®

Source: PDB ID: 1HGUChantalat, L. Jones, N.D., Korber, F., Navaza, J., Pavlovsky, A.G. (1995) The crystal-structure of wild-type growth-hormone at 2.5 Angstrom resolution. Protein Pept. Lett. 2: 333-340


Analytical methods to characterize somatropin in comparability studiesMolecular parameter Method

Edman sequencing

Peptide map with MS detection

Mass Mass spectrometry: MALDI-TOF, ESI-MS

Capillary electrophoresisCharge

Immunological tests Immunoblotting

Circular dichroism spectroscopy

1D {1H} NMR spectroscopy

Reversed phase chromatography

Isoelectric focusing

Ion exchange chromatography

Size exclusion chromatographySize

Gel electrophoresis

In-vivo bioassays

Cell proliferation assay

Primary structure

Spatial structure(secondary and tertiary)

Polarity

Biological activity


Assessment of identity: Complete sequence coverage by combination of peptide maps

AA# 1 2 3 4 5 1 0 0 0 0 0 T1 T2 T4 T6 FPTIPLSRLFDNAMLRAHRLHQLAFDTYQEFEEAYIPKEQKYSFLQNPQT G1 G2 G3 G5 G6 AA# 1 5 6 7 8 9 0 1 0 0 0 0 0 T7 T8 T9 T10 SLCFSESIPTPSNREETQQKSNLELLRISLLLIQSWLEPVQFLRSVFANS G7 G8 1 1 1 1 1 1 0 1 2 3 4 5 1 0 0 0 0 0 T11 T12 T13 T14 T15 LVYGASDSNVYDLLKDLEEGIQTLMGRLEDGSPRTGQIFKQTYSKFDTNS G13 AA# 1 1 1 1 1 5 6 7 8 9 1 0 0 0 0 T16 T18 T19 T20 T19 HNDDALLKNYGLLYCFRKDMDKVETFLRIVQCRSVEGSCGF G18 G21 G22

Complete sequence coverage is obtained by combination of the detected peptides from digests with trypsin (blue) and Glu-C (orange).


Considerations for analytical methods:Impact of matrix and sample preparation

Sample preparation and analytical methods must be developed to account for differences in sample matrices.

40 45 50 55 60

04-WHOOmnitrope

T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4

40 45 50 55 60

04-WHOOmnitrope

T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4Product 1

40 45 50 55 60

04-WHOOmnitrope

T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4

40 45 50 55 60

04-WHOOmnitrope

T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4Product 1

Digestion without buffer exchange

•Somatropin NIBSC 98/574•Somatropin, liquid formulation, pH 6.2

40 45 50 55 60

04-WHOOmnitrope, buffer exchange

T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4

40 45 50 55 60


T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4

Product 1, buffer exchange

40 45 50 55 60


T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4

40 45 50 55 60


T4+T

5

T10

(T6+

T7)-S

-S-T

16T6

-S-S

-T16

T1-1 T1

0-2

T11

T4

Product 1, buffer exchange

Digestion with buffer exchange

•Somatropin NIBSC 98/574•Somatropin, liquid formulation, after buffer exchange

Comparability studiesScope, common elements, differences


Comparability studies in the development of Omnitrope®

• Biosimilarity: Comparison to a reference product• Process changes: Comparison between a pre- and post-change

product• Novel formulations: Comparison of different drug product

formulations, and assessment of potential differences in the profile of product-related variants

Scope


Comparability studies in the development of Omnitrope®

• Science and knowledge based approach− Previous knowledge− Risk assessment with respect to potential impact of changes/

differences− Identification of Critical Quality Attributes (CQA), Critical Process

Parameters (CPP)• Focus on analytical methods to monitor CQA’s, CPP’s• Type and extend of studies (physicochemical, biological,

preclinical, clinical) is determined in a step-wise approach based on these consideration and the knowledge regarding product and process.

Common approach


Comparability studies in the development of Omnitrope

• Setting acceptance criteria:− Considering the previous experience with the product at the

biosimilar developer and the information in the public domain− Biosimilarity: Considering the batch-to-batch variability of the

reference product− Process changes: Based on the historical performance of the

process • Differences need to be assessed in appropriate studies and

scientifically justified to exclude an impact on efficacy and safety of the product

− Classification of the product variants into product-related substances or impurities (ICH Q6B)

Execution

Comparability studiesBiosimilarity


In a representative indication (growth hormone deficient children)

Comprehensive analysis of protein

Comparative PK/PD in healthy volunteers

14- day repeat toxicity (rats)local tolerance (rabbits)

In-vitro and in-vivo bioassays

Development according to relevant guidelines

Com

para

bilit

y

Clinical efficacy and safety

PK/PD

Preclinical

Biologicalcharacterization

Physicochemical characterization

Comparability of Omnitrope® with the reference product was established at all stages

Complete stand-alone product and target-directed process development


Controlling the product profile of somatropin:Detecting potential product-related variants

Variant Polarity (RP-HPLC)

Charge (CZE, IEF, IEX)

Size(SEC)

Deamidated X X -

-

-

-

-

-

Thioether - - - X

Aggregates –Covalent dimer

X - X X

X

X

Succinimide X X -

Isoaspartate X - x

Oxidized X - X

Truncated and clipped variants

X X X

Trisulfide X - X

Non-covalent aggregates – Dimer, oligomers

- - -

Peptide mapping

X…resolved and detected; -…not resolvable with this method


A thioether variant of somatropin:Background

Disulfide bridges in somatropin:• Cys(53)-Cys(165)• Cys(182)-Cys(189)

Thioether variant:• Cys(182)-Cys(189)*

* Datola, A. et al., ChemMedChem, 2 (2007) 1181-1189.

IVQCRSS

SVEGSCGF

IVQCRS

SVEGSCGF


0

20

40

60

80

100

Rel

ativ

e A

bund

ance

684.83

685.34

685.84

678 680 682 684 686 688 690 692 694 696 698 700 702 704 706m/z

0

20

40

60

80

100 700.82

701.32

701.82

702.32

A thioether variant of somatropin:Detection and identification

Time (min)17 18 19 20 21

Reference product

Omnitrope

Somatropin X

T20-

SS-T

21

T20-

S-T2

1

T20-S-S-T21, (M+2H)2+

Expected mass: 700.82Found mass: 700.82

Detection by peptide mapping Identification by high resolution mass spectrometry

T20-S-T21, (M+2H)2+

Expected mass: 684.83Found mass: 684.83


Phase I PK/PD study program

Protocol Design Objective No. Subj. Dose

EP2K99PhI S USA Double-blind, randomised, 2-way cross-over, placebo-controlled study

Phase I safety study in healthy volunteers to assess the safety, tolerance and pharmacokinetics of Omnitrope lyophilised powder (API Covance).Feasibility study to demonstrate that continuous i.v infusion of octreotide is effective to suppress endogenous GH secretion in healthy subjects

12 5 mg single dose s.c. injection

EP2K99PhI USA Double-blind, randomised, 2-way cross-over, controlled study

Phase I study in healthy volunteers to compare the pharmacokinetics, pharmacodynamics and safety between Omnitrope lyophilised powder (API Covance) and Genotropin


EP2K00PhI AQ Double-blind, randomised, 2-way cross-over, controlled study

Phase I study in healthy volunteers to compare the pharmacokinetics, pharmacodynamics and safety between Omnitrope lyophilised powder (API Covance) and Omnitrope 3.3 mg/ml solution f.i. (API Kundl)


EP00-104 Double-blind, randomised, 3-way cross-over, controlled study

Phase I study in healthy volunteers to compare the pharmacokinetics, pharmacodynamics and safety between Omnitrope 5mg/ml powder f.s.f.i., Omnitrope 3.3 mg/ml solution f.i. and Genotropin


EP00-105 Double-blind, randomised, 3-way cross-over, controlled study

Phase I study in healthy volunteers to compare the pharmacokinetics, pharmacodynamics and safety between Omnitrope 5mg/ml powder f.s.f.i., Omnitrope 6.7 mg/ml solution f.i. and Genotropin



Phase III Clinical Program to confirm comparable safety and efficacy

Protocol Design Objective No. Subj. Dose

EP2K99PhIII

EP2K00PhIIIFo

EP2K00PhIIIAQ part A

EP2K00PhIIIAQ part B

EP2K02PhIIILyo

Randomized, controlled, open, multicenter study in GHD children

Efficacy and safety -Demonstrate therapeutic equivalence between Omnitrope Lyophilized Powder and Genotropin

89 Omnitrope Lyophilized Powder vs. Genotropin0.03 mg/kg daily, sc

Controlled, open, multicenter study in GHD children

Efficacy and safety -Follow-up study to demonstrate therapeutic equivalence between Omnitrope Lyophilized Powder and Genotropin

86 Omnitrope Lyophilized Powder vs. Genotropin0.03 mg/kg daily, sc

Controlled, open, multicenter study in GHD children

Efficacy and safety -Demonstrate therapeutic equivalence between Omnitrope Lyophilized Powder and Omnitrope Liquid Formulation

86 Omnitrope Lyophilized Powder vs. Omnitrope Liquid Formulation0.03 mg/kg daily, sc

Open, multicenter study in GHD children

Demonstrate long term safety and efficacy of Omnitrope Liquid Formulation

86 Omnitrope Liquid Formulation0.03 mg/kg daily, sc

Pivotal safety study

Open, multicenter study in GHD children

Safety and efficacy -Confirmation of low immunogenicity of Omnitrope Lyophilized Powder

51 Omnitrope Lyophilized Powder 0.03 mg/kg daily, sc

Pivotal efficacy study


Biosimilarity: Comparability with the reference product

• A biosimilar product is designed to meet the criteria of the reference product with regards to quality, safety and efficacy.

• Biosimilars development requires complete product and process development plus comparative testing at all relevant levels.

• The comparability of Omnitrope® to the reference product was shown using a comprehensive array of test methods.

• Pre-clinical and clinical assessment confirmed similarity and clinical equivalence.

Targeted development/

Quality by Design

Quality

Module 3

Physico-chemical and

biological comparability

Comparative preclinical

studies

Comparative clinical studies

Risk Management

Plan

Comparative

Reference product

Biosimilar product

Comparative

Comparability studiesProcess changes


Comparability assessment for process changes

• Acceptance criteria for comparability: − Based on the historical performance of the process (batch

history) and understanding of the product.− Data from reference product may be used as supportive data in

assessment of the change• Approaches:

− Submission of the entire comparability study− Submission of a comparability protocol ahead of implementation

of changes• Extend of the studies:

− Varies with extend of change− For scale or site changes to a well-controlled and understood

process physicochemical and biological data were found appropriate and sufficient

Comparability studiesNovel formulations


Liquid formulations of somatropin

Stable liquid formulation, containing buffer system, tonicity modifying excipients, surface active excipients, antimicrobial preservation agent

Lyophilized and liquid formulation after submission to mechanical stress:Particle profile in the subvisible range

Particle count512 channels

0.5 µm – 450 µm

Dynamic Light Scattering

0.1 nm – 1 µm

0

5

10

15

0.1 1 10 100 1000

Inte

nsity

(%)

Size (d.nm)

Size Distribution by Intensity

0

5

10

15

20

0.1 1 10 100 1000

Inte

nsity

(%)

Size (d.nm)

Size Distribution by Intensity

Reconstituted powder Liquid formulation


Qualification of a liquid formulation of Omnitrope®

Efficacy and safety of profile of product-related variants confirmed in Phase III studies

Equivalence to lyophilized formulation and reference product in Phase I studies

Toxicology and local tolerance studies, potency in animal model of fresh and aged product

Biological characterization of aged product and product-related variants

Stability studies;characterization of aged product and of product-related variants at end of shelf life

Biological characterization

Preclinical

PK/PD

Clinical safety & efficacy

Physicochemical characterization


Conclusions

• Assessments of comparability are crucial components in the development of any biopharmaceutical product, and specifically so for biosimilar development.

• The extend of a comparability study needs to be based on the magnitude of the introduced change and the knowledge of product and process.

• A set of orthogonal analytical tools is essential to overcome the limitations of single methods and to obtain a complete picture of the quality of the product – biosimilar candidate and reference product.

• The efficacy and safety and the comparability of Omnitrope® with the reference product were shown combining data from physico-chemical, biological, preclinical and clinical studies.

• Data from all these levels were used to demonstrate that the liquid formulations of Omnitrope® have consistent quality characteristics, and are efficacious and safe formulations of somatropin.


Acknowledgements

Technical team• Sabine Fürtinger

• Bernt Pragl

• Hansjörg Toll

Development concepts and registration• Ajaz Hussain

• Martin Schiestl

• Ingrid Schwarzenberger

• Jörg Windisch

Documents

Comparability assessment in the development of follow …c.ymcdn.com/sites/ · 6 Comparability assessment in the development of follow-on ... 678 680 682 684 686 688 690 692 694 696